Comparison of Two Modern Survival Prediction Tools, SORG-MLA and METSSS, in Patients With Symptomatic Long-bone Metastases Who Underwent Local Treatment With Surgery Followed by Radiotherapy and With Radiotherapy Alone.

BACKGROUND: Survival estimation for patients with symptomatic skeletal metastases ideally should be made before a type of local treatment has already been determined. Currently available survival prediction tools, however, were generated using data from patients treated either operatively or with local radiation alone, raising concerns about whether they would generalize well to all patients presenting for assessment. The Skeletal Oncology Research Group machine-learning algorithm (SORG-MLA), trained with institution-based data of surgically treated patients, and the Metastases location, Elderly, Tumor primary, Sex, Sickness/comorbidity, and Site of radiotherapy model (METSSS), trained with registry-based data of patients treated with radiotherapy alone, are two of the most recently developed survival prediction models, but they have not been tested on patients whose local treatment strategy is not yet decided. QUESTIONS/PURPOSES: (1) Which of these two survival prediction models performed better in a mixed cohort made up both of patients who received local treatment with surgery followed by radiotherapy and who had radiation alone for symptomatic bone metastases? (2) Which model performed better among patients whose local treatment consisted of only palliative radiotherapy? (3) Are laboratory values used by SORG-MLA, which are not included in METSSS, independently associated with survival after controlling for predictions made by METSSS? METHODS: Between 2010 and 2018, we provided local treatment for 2113 adult patients with skeletal metastases in the extremities at an urban tertiary referral academic medical center using one of two strategies: (1) surgery followed by postoperative radiotherapy or (2) palliative radiotherapy alone. Every patient's survivorship status was ascertained either by their medical records or the national death registry from the Taiwanese National Health Insurance Administration. After applying a priori designated exclusion criteria, 91% (1920) were analyzed here. Among them, 48% (920) of the patients were female, and the median (IQR) age was 62 years (53 to 70 years). Lung was the most common primary tumor site (41% [782]), and 59% (1128) of patients had other skeletal metastases in addition to the treated lesion(s). In general, the indications for surgery were the presence of a complete pathologic fracture or an impending pathologic fracture, defined as having a Mirels score of ≥ 9, in patients with an American Society of Anesthesiologists (ASA) classification of less than or equal to IV and who were considered fit for surgery. The indications for radiotherapy were relief of pain, local tumor control, prevention of skeletal-related events, and any combination of the above. In all, 84% (1610) of the patients received palliative radiotherapy alone as local treatment for the target lesion(s), and 16% (310) underwent surgery followed by postoperative radiotherapy. Neither METSSS nor SORG-MLA was used at the point of care to aid clinical decision-making during the treatment period. Survival was retrospectively estimated by these two models to test their potential for providing survival probabilities. We first compared SORG to METSSS in the entire population. Then, we repeated the comparison in patients who received local treatment with palliative radiation alone. We assessed model performance by area under the receiver operating characteristic curve (AUROC), calibration analysis, Brier score, and decision curve analysis (DCA). The AUROC measures discrimination, which is the ability to distinguish patients with the event of interest (such as death at a particular time point) from those without. AUROC typically ranges from 0.5 to 1.0, with 0.5 indicating random guessing and 1.0 a perfect prediction, and in general, an AUROC of ≥ 0.7 indicates adequate discrimination for clinical use. Calibration refers to the agreement between the predicted outcomes (in this case, survival probabilities) and the actual outcomes, with a perfect calibration curve having an intercept of 0 and a slope of 1. A positive intercept indicates that the actual survival is generally underestimated by the prediction model, and a negative intercept suggests the opposite (overestimation). When comparing models, an intercept closer to 0 typically indicates better calibration. Calibration can also be summarized as log(O:E), the logarithm scale of the ratio of observed (O) to expected (E) survivors. A log(O:E) > 0 signals an underestimation (the observed survival is greater than the predicted survival); and a log(O:E) < 0 indicates the opposite (the observed survival is lower than the predicted survival). A model with a log(O:E) closer to 0 is generally considered better calibrated. The Brier score is the mean squared difference between the model predictions and the observed outcomes, and it ranges from 0 (best prediction) to 1 (worst prediction). The Brier score captures both discrimination and calibration, and it is considered a measure of overall model performance. In Brier score analysis, the "null model" assigns a predicted probability equal to the prevalence of the outcome and represents a model that adds no new information. A prediction model should achieve a Brier score at least lower than the null-model Brier score to be considered as useful. The DCA was developed as a method to determine whether using a model to inform treatment decisions would do more good than harm. It plots the net benefit of making decisions based on the model's predictions across all possible risk thresholds (or cost-to-benefit ratios) in relation to the two default strategies of treating all or no patients. The care provider can decide on an acceptable risk threshold for the proposed treatment in an individual and assess the corresponding net benefit to determine whether consulting with the model is superior to adopting the default strategies. Finally, we examined whether laboratory data, which were not included in the METSSS model, would have been independently associated with survival after controlling for the METSSS model's predictions by using the multivariable logistic and Cox proportional hazards regression analyses. RESULTS: Between the two models, only SORG-MLA achieved adequate discrimination (an AUROC of > 0.7) in the entire cohort (of patients treated operatively or with radiation alone) and in the subgroup of patients treated with palliative radiotherapy alone. SORG-MLA outperformed METSSS by a wide margin on discrimination, calibration, and Brier score analyses in not only the entire cohort but also the subgroup of patients whose local treatment consisted of radiotherapy alone. In both the entire cohort and the subgroup, DCA demonstrated that SORG-MLA provided more net benefit compared with the two default strategies (of treating all or no patients) and compared with METSSS when risk thresholds ranged from 0.2 to 0.9 at both 90 days and 1 year, indicating that using SORG-MLA as a decision-making aid was beneficial when a patient's individualized risk threshold for opting for treatment was 0.2 to 0.9. Higher albumin, lower alkaline phosphatase, lower calcium, higher hemoglobin, lower international normalized ratio, higher lymphocytes, lower neutrophils, lower neutrophil-to-lymphocyte ratio, lower platelet-to-lymphocyte ratio, higher sodium, and lower white blood cells were independently associated with better 1-year and overall survival after adjusting for the predictions made by METSSS. CONCLUSION: Based on these discoveries, clinicians might choose to consult SORG-MLA instead of METSSS for survival estimation in patients with long-bone metastases presenting for evaluation of local treatment. Basing a treatment decision on the predictions of SORG-MLA could be beneficial when a patient's individualized risk threshold for opting to undergo a particular treatment strategy ranged from 0.2 to 0.9. Future studies might investigate relevant laboratory items when constructing or refining a survival estimation model because these data demonstrated prognostic value independent of the predictions of the METSSS model, and future studies might also seek to keep these models up to date using data from diverse, contemporary patients undergoing both modern operative and nonoperative treatments. LEVEL OF EVIDENCE: Level III, diagnostic study.

Enzymatic denaturation versus excessive fatigue loading degeneration: Effects on the time-dependent response of the intervertebral disc.

Degenerative disc disease (DDD), regardless of its phenotype and clinical grade, is widely associated with low back pain (LBP), which remains the single leading cause of disability worldwide. This work provides a quantitative methodology for comparatively investigating artificial IVD degeneration via two popular approaches: enzymatic denaturation and fatigue loading. An in-vitro animal study was used to study the time-dependent responses of forty fresh juvenile porcine thoracic IVDs in conjunction with inverse and forward finite element (FE) simulations. The IVDs were dissected from 6-month-old-juvenile pigs and equally assigned to 5 groups (intact, denatured, low-level, medium-level, high-level fatigue loading). Upon preloading, a sinusoid cyclic load (Peak-to-peak/0.1-to-0.8 MPa) was applied (0.01-10 Hz), and dynamic-mechanical-analyses (DMA) was performed. The DMA outcomes were integrated with a robust meta-model analysis to quantify the poroelastic IVD characteristics, while specimen-specific FE models were developed to study the detailed responses. The results demonstrated that enzymatic denaturation had a more significantly pronounced effect on the resistive strength and shock attenuation capabilities of the intervertebral discs. This can be attributed to the simultaneous disruption of the collagen fibers and water-proteoglycan bonds induced by trypsin digestion. Fatigue loading, on the other hand, primarily influenced the disc's resistance to deformation in a frequency-dependent pattern, where alterations were most noticeable at low loading frequencies. This study confirms the intricate interplay between the biochemical changes induced by enzymatic processes and the mechanical behavior stemming from fatigue loading, suggesting the need for a comprehensive approach to closely mimic the interrelated multifaceted processes of human disc degeneration.

Effects of pilates exercises on radiographic lumbo-pelvic alignment and range of motion in non-specific low back pain patients.

OBJECTIVES: To determine the effects of Pilates exercises on lumbo-pelvic alignment in non-specific low back pain (NSLBP) patients. METHODS: Twenty-two patients (Male:7; Female:15) with NSLBP aged 20-65 years were recruited and classified based on a modified O'Sullivan's classification system into flexion pattern (FP) or active extension pattern (EP) groups. Oswestry Disability index (ODI), Roland-Morris Disability Questionnaire (RMDQ) as well as radiographic lumbar global range of motion (ROM) and lumbo-pelvic alignment were measured Pre- and immediately post-intervention and at 6- and 12-month. The intervention included supervised six-weeks Pilates program with 60 min per session and up to two sessions per week. RESULTS: Lumbar lordosis, sacral slope and sacral inclination were found to be significantly different between the FP and EP groups based on the O'Sullivan's classification system. However, despite the significant changes in RMDQ (p = 0.001), no significant changes were found for any of the alignment parameters (p > 0.05) post intervention for both groups. For lumbar global ROM, a statistically significant change was observed for the EP group (p = 0.028) but not for the FP group (p = 0.249). No significant correlations were identified between any of the self-reported outcomes, radiographic alignment and ROM parameters. CONCLUSIONS: Patients self-perceived long-term functional improvements based on responding to questionnaires after Pilates exercises were not reflected in significant changes in lumbo-pelvic alignment or lumbar ROM. This may be due to the current cohort demonstrating within normal ranges due to the lesser severity of their condition, but further research is needed for clarification.

ASIC3 roles in mechanosensitive elongation of nucleus pulposus cells.

Morphological changes of the nucleus pulposus (NP) cells occur concomitantly as part of the intervertebral disc (IVD) degeneration and excessive mechanical loading has been speculated as a significant key factor for contributing to such morphological changes. Therefore, we hypothesize that stress exerted on NP cells can cause a deformity of nucleus in response. The changes of cell morphology is observed in degenerative nucleus pulposus. One of the reasons for degeneration of NP is due to overloading of NP especially in the obese population. So the nucleus deformity caused by stress/force is of our study interest. To delineate the effects and role of mechanical stress, we developed a 3D assay using hydrogel cultures with a circular hole generated with needle indentation to simulate a local stress concentration along the edge of the hole. A stressed zone, encompassing 100 µm of range from the circular edge, is defined based on stress concentration calculation to enable quantitative analysis against the control zone. Our results demonstrated that the circular hole produces stress-induced morphological changes in NP cells. The tangential elongation of NP cells and their nucleus shape changes in the stressed zone are significantly increased compared to the non-stressed control zone. It is proposed that the cell elongation is a direct response to elevated stress within the stressed zone. Subsequently we found the stress induced morphological changes of the NP cells can be significantly reduced by inhibiting ASIC3. This suggests ASIC3 plays an important role of play in mechano-signaling of NP cells.

Involvement of ASIC3 and Substance P in Therapeutic Ultrasound-Mediated Analgesia in Mouse Models of Fibromyalgia.

Therapeutic ultrasound (tUS) is widely used in chronic muscle pain control. However, its analgesic molecular mechanism is still not known. Our objective is to reveal the mechanism of the tUS-induced analgesia in mouse models of fibromyalgia. We applied tUS in mice that have developed chronic hyperalgesia induced by intramuscular acidification and determined the tUS frequency at 3 MHz, dosage at 1 W/cm2 (measured output as 6.3 mW/cm2) and 100% duty cycle for 3 minutes having the best analgesic effect. Pharmacological and genetic approaches were used to probe the molecular determinants involved in tUS-mediated analgesia. A second mouse model of fibromyalgia induced by intermittent cold stress was further used to validate the mechanism underlying the tUS-mediated analgesia. The tUS-mediated analgesia was abolished by a pretreatment of NK1 receptor antagonist-RP-67580 or knockout of substance P (Tac1-/-). Besides, the tUS-mediated analgesia was abolished by ASIC3-selective antagonist APETx2 but not TRPV1-selective antagonist capsazepine, suggesting a role for ASIC3. Moreover, the tUS-mediated analgesia was attenuated by ASIC3-selective nonsteroid anti-inflammation drugs (NSAIDs)-aspirin and diclofenac but not by ASIC1a-selective ibuprofen. We next validated the antinociceptive role of substance P signaling in the model induced by intermittent cold stress, in which tUS-mediated analgesia was abolished in mice lacking substance P, NK1R, Asic1a, Asic2b, or Asic3 gene. tUS treatment could activate ASIC3-containing channels in muscle afferents to release substance P intramuscularly and exert an analgesic effect in mouse models of fibromyalgia. NSAIDs should be cautiously used or avoided in the tUS treatment. PERSPECTIVE: Therapeutic ultrasound showed analgesic effects against chronic mechanical hyperalgesia in the mouse model of fibromyalgia through the signaling pathways involving substance P and ASIC3-containing ion channels in muscle afferents. NSAIDs should be cautiously used during tUS treatment.

On minimal focal distance of a focused ultrasound probe for neuromodulation.

Focal distance is a key parameter for a focused ultrasound probe, especially in mouse brain stimulation where targets are right below the skull. A closed-form solution for the minimal focal distance with a given transducer size was derived in this study to facilitate precise focal spot alignment with targets in the mouse brain. The spherical profile corresponding to the minimal focal distance does not produce accurate focusing. An iterative algorithm based on Snell's law was introduced for lens profile calculation. With a suitable step size, an accurate lens profile can be obtained for the minimal focal distance.

Activation of Mechanosensitive Ion Channels by Ultrasound.

Mechanosensitive channels (MSCs) play an important role in how cells transduce mechanical stimuli into electrical or chemical signals, which provides an interventional possibility through the manipulation of ion channel activation using different mechanical stimulation conditions. With good spatial resolution and depth of penetration, ultrasound is often proposed as the tool of choice for such therapeutic applications. Despite the identification of many ion channels as mechanosensitive in recent years, only a limited number of MSCs have been reported to be activated by ultrasound with substantial evidence. Furthermore, although many therapeutic implications using ultrasound have been explored, few offered insights into the molecular basis and the biological effects induced by ultrasound in relieving pain and accelerate tissue healing. In this review, we examined the literature, in particular studies that provided evidence of cellular responses to ultrasound, with and without the target ion channels. The ultrasound activation conditions were then summarized for these ion channels, and these conditions were related to their mode of activation based on the current biological concepts. The overall goal is to bridge the results relating to the activation of MSCs that is specific for ultrasound with the current knowledge in molecular structure and the available physiological evidence that may have facilitated such phenomena. We discussed how collating the information revealed by available scientific investigations helps in the design of a more effective stimulus device for the proposed translational purposes. Traditionally, studies on the effects of ultrasound have focused largely on its mechanical and physical interaction with the targeted tissue through thermal-based therapies as well as non-thermal mechanisms including ultrasonic cavitation; gas body activation; the direct action of the compressional, tensile and shear stresses; radiation force; and acoustic streaming. However, the current review explores and attempts to establish whether the application of low-intensity ultrasound may be associated with the activation of specific MSCs, which in turn triggers relevant cell signaling as its molecular mechanism in achieving the desired therapeutic effects. Non-invasive brain stimulation has recently become an area of intense research interest for rehabilitation, and the implication of low-intensity ultrasound is particularly critical given the need to minimize heat generation to preserve tissue integrity for such applications.

Piezoelectric effect stimulates the rearrangement of chondrogenic cells and alters ciliary orientation via atypical PKCζ.

Therapeutic ultrasound was administered to patients suffering from bone fracture with FDA approval. Bone and cartilage are piezoelectric materials. To investigate the effects of piezoelectricity on the cells of chondrogenic lineage, we applied ultrasound stimulation on an AT-cut quartz coverslip to generate electric field fluctuations. The bone-marrow-derived mesenchymal stem cells (BMMSC) and primary chondrocytes were cultured on either glass or quartz coverslips for ultrasound stimulation. The cells were immunofluorescent-labeled for the assessment of cell arrangement and ciliary orientation. Ultrasound and piezoelectricity both stimulate cell migration and disrupt ciliary orientation induced by directional migration. In particular, piezoelectric effects on cell rearrangement can be abolished by the inhibitor specifically targeting atypical Protein kinase C zeta (PKCζ). Our findings shed light on the possibility of cellular modulation by using piezoelectric manipulation.

Auditory independent low-intensity ultrasound stimulation of mouse brain is associated with neuronal ERK phosphorylation and an increase of Tbr2 marked neuroprogenitors.

Transcranial ultrasound stimulation is an emerging technique for the development of a non-invasive neuromodulation device for the treatment of various types of neurodegenerations and brain damages. However, there are very few studies that have quantified the optimal ultrasound dosage and the long-term associated effects of transcranial ultrasound treatments of brain diseases. In this study, we used a simple ex vivo hippocampal tissues stimulated by different dosages of ultrasound in combination with different chemical treatments to quantify the required energy for a measurable effect. After determining the most desirable ex vivo stimulation conditions, it was then replicated for the in vivo mouse brains. It was discovered that transcranial ultrasound promoted the increase of Tbr2-expressing neural progenitors in an ASIC1a-dependent manner. Furthermore, such effect was observable at least a week after the initial ultrasound treatments and was not abolished by auditory toxicity.

Dynamic Pressure Stimulation Upregulates Collagen II and Aggrecan in Nucleus Pulposus Cells Through Calcium Signaling.

STUDY DESIGN: An in vitro study to investigate the effect of pressure stimulation on nucleus pulposus (NP) cells. OBJECTIVE: The aim of this study was to investigate the question whether physical stimulation can be leveraged to enhance extracellular matrix (ECM) synthesis as a preventive measure for intervertebral disc (IVD) degeneration. SUMMARY OF BACKGROUND DATA: ECM plays an important role in regulating hydration and pressure balance of the IVD. METHODS: Cellular stimulation devices with different pressurizing protocols were used to create a pressurized environment to cells cultures. The setup was used to mimic the pressurized conditions within IVD to investigate the effect of pressure stimulation on NP cells. RESULTS: Pressure stimulation at 300 kPa can enhance the synthesis of ECM proteins Collagen II and aggrecan in NP cells and the effect of dynamic pressure stimulation outperformed the static one. The difference between static and dynamic pressure stimulation was due primarily to calcium signaling activated by pressure fluctuation. The superior effect of dynamic pressure holds for a wide range of stimulation durations, relating to the range of spontaneous calcium oscillations in NP cells. CONCLUSION: The results link mechanotransduction to the downstream ECM protein synthesis and suggest slow exercises that correspond with spontaneous calcium oscillations in NP cells can be effective to stimulate ECM synthesis in IVD.

A role for substance P and acid-sensing ion channel 1a in prolotherapy with dextrose-mediated analgesia in a mouse model of chronic muscle pain.

ABSTRACT: Prolotherapy is widely used in pain control and tissue repair in pain medicine. The classical mode is injection with hypertonic dextrose in muscle or perimysium. However, the analgesic mechanism is still not known. Here, we successfully established dextrose-mediated antinociception in a mouse model of fibromyalgia. The antinociceptive effects of dextrose injections were evaluated in a mouse model of fibromyalgia, in which bilateral chronic mechanical hyperalgesia was induced by unilateral intramuscular acid injection. The injectant (dextrose), dose (≥5%), and volume (>10 µL), but not osmolarity, were essential for the prolotherapy. Further studies showed that the activation of acid-sensing ion channel 1a (ASIC1a), neural activation, and the release of substance P from muscle afferents were required in the dextrose-induced reduction of mechanical hypersensitivity. Both pharmacological blockade and genetic deletion of ASIC1a or substance P as well as lidocaine abolished the dextrose-induced antinociception in mice with chronic hyperalgesia. Moreover, intramuscular dextrose injection induced phosphorylated extracellular signal-regulated kinase expression in dorsal root ganglion neurons expressing substance P; the phosphorylated extracellular signal-regulated kinase expression was inhibited by the ASIC1a antagonist PcTx1. The optimal settings for prolotherapy in fibromyalgia-like pain are dextrose dependent and volume dependent, and the peripheral antinociception involves ASIC1a and substance P signaling in muscle afferents. This study suggests a possible mechanism of action of dextrose prolotherapy in noninflammatory muscle pain such as fibromyalgia and provides insights into treating other types of chronic pain.

ASIC1a is required for neuronal activation via low-intensity ultrasound stimulation in mouse brain.

Accumulating evidence has shown transcranial low-intensity ultrasound can be potentially a non-invasive neural modulation tool to treat brain diseases. However, the underlying mechanism remains elusive and the majority of studies on animal models applying rather high-intensity ultrasound that cannot be safely used in humans. Here, we showed low-intensity ultrasound was able to activate neurons in the mouse brain and repeated ultrasound stimulation resulted in adult neurogenesis in specific brain regions. In vitro calcium imaging studies showed that a specific ultrasound stimulation mode, which combined with both ultrasound-induced pressure and acoustic streaming mechanotransduction, is required to activate cultured cortical neurons. ASIC1a and cytoskeletal proteins were involved in the low-intensity ultrasound-mediated mechanotransduction and cultured neuron activation, which was inhibited by ASIC1a blockade and cytoskeleton-modified agents. In contrast, the inhibition of mechanical-sensitive channels involved in bilayer-model mechanotransduction like Piezo or TRP proteins did not repress the ultrasound-mediated neuronal activation as efficiently. The ASIC1a-mediated ultrasound effects in mouse brain such as immediate response of ERK phosphorylation and DCX marked neurogenesis were statistically significantly compromised by ASIC1a gene deletion. Collated data suggest that ASIC1a is the molecular determinant involved in the mechano-signaling of low-intensity ultrasound that modulates neural activation in mouse brain.

Elevation of Intra-Cellular Calcium in Nucleus Pulposus Cells with Micro-Pipette-Guided Ultrasound.

Modulation of intra-cellular calcium by ultrasound offers a possible means for therapeutic applications. One such possibility is the modulation of nucleus pulposus cells as a preventive measure for inter-vertebral disc degeneration. We report a cellular stimulation device (micro-pipette ultrasound) using a glass micro-pipette as a waveguide to deliver ultrasound through the pipette tip and to elevate intra-cellular calcium in nucleus pulposus cells. The device generates two relevant stimuli at the cellular level: ultrasound propagation throughout the cell and acoustic streaming on the apical side. Ultrasound is radiated from a tip of a few microns, and its amplitude is proportional to the input voltage; acoustic streaming can be controlled by the duty factor. The novelty of the device is to impose a unique cellular loading: shear stress on cell apical surfaces combined with compressional waves propagating through the cells. G protein-coupled receptors and acid-sensing ion channel 3 were shown to play a role in calcium elevation by micro-pipette ultrasound in nucleus pulposus cells. Our results demonstrate that micro-pipette ultrasound can be an effective tool to elevate intra-cellular calcium levels in different cells, facilitating the identification of different mechanoreceptors in action.

Gelatin-Poly (γ-Glutamic Acid) Hydrogel as a Potential Adhesive for Repair of Intervertebral Disc Annulus Fibrosus: Evaluation of Cytocompatibility and Degradability.

STUDY DESIGN: An in vitro experimental study testing a Gelatin-poly (γ-glutamic acid) hydrogel for disc repair. OBJECTIVE: To evaluate the cytocompatibility and degradability of the above mentioned hydrogel for intervertebral disc annular fibrosis (AF) repair. SUMMARY OF BACKGROUND DATA: No repair strategies for correcting annular defects in lumbar discectomy have been clinically well recognized. Exogenous supplementation of regenerative materials to fill defects is a minimally invasive way to restore compromised mechanical properties. The injected materials, most commonly gelatin-based materials with cross-linking agents, serve as sealants and as a scaffold for incorporating biomaterials for augmentation. However, cytotoxicity of hydrogel crosslinking agents is of concern in developing viable materials. METHODS: This in vitro experimental study evaluated a newly developed gelatin-based hydrogel for intervertebral disc AF repair. Mechanical strength was augmented by γ-PGA, and 1-(3-dimethylaminopropyl)-3-ethyl-carbodiimide hydrochloride (EDC) was used for material crosslinking. Isolated bovine tail intervertebral discs (IVDs) were used to test the hydrogel, and hydrogel surface monolayer AF cell culture was used to investigate efficacy in hydrogel constructs of different EDC concentrations. Cell metabolic activity was evaluated with Alamar blue assay, cell viability assay with live/dead stain, and sulfated glycosaminoglycan (GAG) and double strain DNA were quantified to evaluate proliferation of implanted cells and synthesis of extracellular matrix (ECM) proteins. RESULTS: EDC concentrations from 10 to 40 mM resulted in significant decreases in AF cell proliferation without obvious influence on cell viability. Higher EDC concentrations resulted in decreased percentage of Alamar blue reduction and GAG and DNA concentration, but did not affect GAG/DNA and live-dead ratios. Degradation tests revealed that higher EDC concentrations decreased the hydrogel degradation rate. CONCLUSION: The developed gelatin-poly (γ-PGA) hydrogel with 20 mM EDC concentration provides an effective gap-filling biomaterial with good cytocompatibility, suggesting substantial promise for use as a sealant for small AF defects.Level of Evidence: N/A.

Low intensity ultrasound enhances cisplatin uptake in vitro by cochlear hair cells.

Drug delivery to the inner ear has been challenging due to the blood-labyrinth barrier. Intracochlear drug delivery is an invasive alternative with less pharmacokinetic variables. In this study, the effect of low intensity ultrasound on drug uptake by hair cells is investigated. Cochlear explants harvested from newborn mice were cultured in a medium containing cisplatin to emulate drug delivered to the endolymph. The results demonstrated the exposure to ultrasound stimulation effectively enhanced cisplatin uptake by hair cells. The uptake started from the apical side of the hair cells and progressed inward as the exposure time increased.

The responses of nucleus pulposus cells to pressure and ultrasound stimulation.

A cellular stimulation device with a pressurized chamber is developed to investigate the effect of ultrasound and pressure fluctuation on nucleus pulposus (NP) cells. The pressurized chamber is designed to emulate the in vivo environment of intervertebral discs, which are under dynamic pressure, and to emulate impact during sports and exercise. Both hydrostatic pressure and ultrasound stimulation increase phosphorylation of ERK (pERK) in NP cells, and promote its translocation into nucleus. This increase in pERK levels might be activated through calcium signaling pathways as intracellular calcium in NP cells was strongly elevated by pressure changes.

Piezoelectric stimulation by ultrasound facilitates chondrogenesis of mesenchymal stem cells.

A cellular stimulation device utilizing an AT-cut quartz coverslip mounted on an ultrasonic live imaging chamber is developed to investigate the effect of piezoelectric stimulation. Two types of chambers deliver ultrasound at intensities ranging from 1 to 20 mW/cm2 to mesenchymal stem cells (MSCs) seeded on the quartz coverslip. The quartz coverslip imposes additionally localized electric charges as it vibrates with the stimulation. The device was applied to explore whether piezoelectric stimulation can facilitate chondrogenesis of MSCs. The results suggest piezoelectric stimulation drove clustering of MSCs and consequently facilitated chondrogenesis of MSCs without the use of differentiation media.

Upright Balance Control in Individuals with Cervical Myelopathy Following Cervical Decompression Surgery: A Prospective Cohort Study.

Patients with cervical myelopathy may manifest impairments in functional activities and balance control caused by compression of the spinal cord. The objective of the current study was to determine long-term changes in the upright balance control of patients with cervical myelopathy who had undergone cervical decompression surgery. This is a prospective cohort study from the preoperative phase to 3 months, 6 months, and 1 year postsurgery. Fifty-three patients with cervical myelopathy were recruited for the cervical myelopathy group and 22 age-matched healthy controls were recruited for the control group. Functional assessments including Japanese Orthopedic Association Cervical Myelopathy Evaluation Questionnaire-Lower Extremity Function (JOACMEQ-LEF) and 10-second step test; as well as balance assessments including postural sway (center-of-pressure: COP) were performed for both groups. The JOACMEQ-LEF (p = 0.036) scores of the myelopathy group improved postoperatively, and a significant decrease in COP variables of postural sway was observed. The upright posture was less stable in the myelopathy group than in the control group (p < 0.05) both before and after surgery. The effect size and standard response mean of the COP variables ranged from -0.49 to 0.03 at 3 months, 6 months, and 1 year postsurgery. The upright balance control had improved significantly 6 months after decompression surgery. However, the balance control of the patients who had undergone decompression surgery remained less stable than that of the age-matched healthy controls. Balance training should be initiated before 6 months postsurgery to accelerate balance control recovery in patients with cervical myelopathy.

Low Intensity Ultrasound Induces Epithelial Cell Adhesion Responses.

In this study, we investigated the cellular mechanosensitive responses to a low intensity ultrasound (LIUS) stimulation (ISATA = 1 mW/cm2, pressure = 10 kPa). The dose and temporal effects at cell-substrate adhesion (CSA) at the basal level and cell-cell adhesion (CCA) at the apical level are reported in detail. A model of mouse mammary gland epithelial cells (EpH4) and the phosphorylation of mechanosensitive 130 kDa Crk-associated substrate (p130CAS) as an indicator for cellular responses were used. The intensity of phospho-p130CAS was found to be dependent on LIUS stress level, and the p130CAS was phosphorylated after 1 min stimulation at CSA. The phospho-p130CAS was also found to increase significantly at CCA upon LIUS stimulation. We confirmed that the cellular responses to ultrasound are immediate and dose dependent. Ultrasound affects not only CSA but also CCA. An E-cadherin knockout (EpH4ECad-/-) model also confirmed that phosphorylation of p130CAS at CCA is related to E-cadherins.

Perturbation-Based Balance Training in Postoperative Individuals With Degenerative Cervical Myelopathy.

Degenerative cervical myelopathy (DCM) is a common aging condition caused by spinal cord compression. Individuals with DCM often presented with residual balance and functional impairments postoperatively. Perturbation-based balance training (PBT) has been shown to have positive effects on populations with neurological disorders but has yet to be investigated in DCM. The objective of this study was therefore to evaluate the effects of PBT on balance and functional performance in postoperative individuals with DCM. Fifteen postoperative individuals with DCM (DCM group) and 14 healthy adults (healthy control group) were recruited. The DCM group received a 4-weeks PBT using a perturbation treadmill. The outcome measures included mean velocity of center of pressure (COP) during quiet standing; center of mass (COM) variance and reaction time to balance perturbation during standing with forward and backward perturbation; gait speed during level ground walking; Timed Up and Go Test (TUG) and disability questionnaire scores including Visual Analog Scale, Neck Disability Index, and Lower Extremity Function of Japanese Orthopaedic Association Cervical Myelopathy Evaluation Questionnaire. The assessments were conducted pre- and post-training postoperatively for the DCM group but only once for the healthy control group. Significant improvements were observed in the mean velocity of COP, COM variance, reaction time, gait speed, and TUG in the DCM group. Disability questionnaire scores were not significantly different after training in DCM group. For between-group comparisons, significant differences that were observed pre-training were not observed post-training. The 4-weeks PBT is a potential rehabilitation strategy for addressing balance and functional impairment in postoperative individuals with DCM. In addition, the post-training performance in the DCM group exhibited trends comparable to those of age-matched healthy controls. Furthermore, the training regimens offer a practical reference for future studies on populations with balance disorders. Future studies complemented with neurophysiological assessments could reveal more information of the underlying mechanisms of PBT.